The respiratory tract is commonly exposed to a variety of oxidizing air pollutants, such as ozone, nitrogen dioxide or cigarette smoke, which can induce inflammatory responses in the respiratory tract, and result in lung injury. Because of its high reactivity, inhaled ozone is expected to interact primarily with respiratory tract lining fluids (RTLFs), the first biological matrix to come into contact with inhaled pollutants. The RTLFs contain significant amounts of low-molecular mass antioxidants (ascorbate, urate, glutathione (GSH) and a-tocopherol), that may serve to protect functional constituents within these RTLFs (e.g. surfactant) as well as the underlying epithelium from direct oxidative injury by inhaled air pollutants. However, the sources and actual concentrations of these RTLF antioxidants are not clear. The applicant hypothesizes that respiratory tract epithelial cells contribute to maintain RTLF low-molecular mass antioxidants and may be involved in antioxidant recycling. The first aim will analyze RTLFs from various regions of the respiratory tract for these low-molecular mass antioxidants, by collecting RTLFs from human volunteers using various nasal and bronchial lavage techniques. The second aim will characterize mechanisms by which the respiratory tract epithelium may contribute to maintain extracellular antioxidant levels, with the use of cultured bronchial and alveolar epithelial cells, and will investigate the importance of such mechanisms in the defense against epithelial injury by inhaled oxidants such as ozone. An important RTLF component, especially during respiratory tract inflammation, is nitric oxide which is known to react readily with ozone. Hence, toxicity due to ozone inhalation is likely to be modulated by increased NO production. Reaction of NO, or its metabolite nitrite, with ozone may serve as a protective mechanism against ozone injury, but may result in formation of reactive nitrogen species (NO2 or ONOO-) that can contribute to ozone toxicity. The third aim is to investigate these possibilities. Finally, as ozone induced lung injury is most likely mediated via initial reaction with RTLF constituents resulting in formation of secondary oxidants, the fourth aim will characterize reaction products with RTLF components, in an attempt to identify secondary toxicants or characteristic oxidation products that can serve as "dosimeters" of in vivo exposure to ozone. This information is likely to increase our understanding of lung injury due to inhalation of air pollutants, and will enable development of strategies to prevent lung injury by inhaled oxidants.